PROJECT SUMMARY Basal-like breast cancer (BLBC) is one of the most aggressive forms of human cancer, because it lacks effective targeted therapies and presents with development of recurrence, distant metastasis, shorter survival, and usually occurs in young women. BLBC has the activated epithelial-mesenchymal transition (EMT) program, which provides cancer cells with the phenotypic and cellular plasticity required for invasion, cancer stem cell (CSC)-like features, and drug resistance. In the last several years, we have systematically studied the role of Snail and Twist, two key EMT-inducers, in BLBC. We discovered that Snail functions to repress genes that prevent EMT (such as E-cadherin), whereas Twist, by interacting with BRD4, serves as a transcription activator to induce genes that enhance EMT (such as Wnt5A). We also discovered that EMT is coupled with a metabolic reprogramming, which provides a metabolic growth advantage for BLBC cells. Our metabolic analyses clearly indicate that BLBC cells also have increased activity for the pathway driving de novo serine synthesis, which is a significant branch derived from glycolysis. The serine biosynthetic pathway is intimately coupled to one-carbon metabolism, which intertwines with the folate and methionine cycles to integrate nutritional inputs from glucose, amino acids and vitamins for the biosynthesis of nucleotides, lipids, and proteins as well as the methylation of chromatin. Our laboratory has discovered that phosphoserine aminotransferase 1 (PSAT1) expression, a key enzyme in serine biosynthesis, is markedly increased in cell lines and clinical BLBC samples. In addition, we found that expression of Twist correlated with the level of PSAT1 in BLBC and that Twist significantly induced PSAT1 expression. Knockdown of PSAT1 expression not only reduced glucose uptake and lactate production but also inhibited cell proliferation, migration and invasion in BLBC. We hypothesize that Twist is the key molecule controlling the expression of PSAT1, which is responsible for the elevated activity of the serine biosynthetic pathway seen in BLBC. Guided by strong preliminary data, we will test this hypothesis by pursuing three specific aims: (1) to elucidate the regulatory mechanism of PSAT1 by Twist in BLBC; (2) to delineate the function of the serine/PSAT1 pathway in BLBC; and (3) to determine the function of the serine/PSAT1 pathway in vivo. Our proposal is innovative and significant, because it incorporates a merged investigation of two essential cancer markers in BLBC, metabolic reprogramming and EMT, and because it leads to the development of an effective precision cancer therapy approach in treating BLBC by combining metabolic drugs with epigenetic inhibitors.